Pro-ACTH/LPH related proteins are present in a variety of different tissues and have been shown to have sites of synthesis outside the pituitary. The expression of this gene in the different tissues may be under different hormonal controls. Using a cloned DNA probe complementary to the pro-ACTH/LPH mRNA, extrapituitary sites of synthesis will be identified by in situ hybridization. The sequence of the pro-ACTH/LPH mRNA in these tissues will be determined using recombinant DNA techniques to determine if more than one gene for the precursor is being expressed in the different tissues. Hormonal regulation of the pro-ACTH/LPH mRNA levels will be monitored by either quantitative in situ hybridization or solution hybridization. Because glucocorticoids are well known regulators of pro-ACTH/LPH in the anterior pituitary but not in other tissues, glucocorticoid receptors will be analyzed in tissues which are expressing the pro-ACTH/LPH gene. Structural and sequence analysis will be performed on the various pro-ACTH/LPH genes isolated from total genomic DNA. The developmental pattern of expression of the pro-ACTH/LPH gene will be monitored using the in situ hybridization techniques. Once sites of synthesis of pro-ACTH/LPH proteins have been identified, the expressing tissues will be cultured with radioactive amino acids and the processing of the precursor analyzed by specific immunoprecipitation and two-dimensional gel electrophoresis. To aid in these studies, monoclonal antibodies to pro-ACTH/LPH related peptides will be generated.